Accurate estimates of DNA quantity are likely to become increasingly important for successful genomic screening of insect populations via recently developed, highly multiplexed genotyping assays and high-throughput se...Accurate estimates of DNA quantity are likely to become increasingly important for successful genomic screening of insect populations via recently developed, highly multiplexed genotyping assays and high-throughput sequencing methods. Here we show that genomic DNA extractions from single Anopheles gambiae Giles using a standard commercial kit-based methodology yield extracts with concentrations below the linear range of spectrophotometric absorbance at 260 nm. Concentrations determined by spectropho- tometry were not reproducible, and are therefore neither accurate nor reliable. However, DNA quantification using a fluorescent nucleic acid stain （PicoGreen） gave highly reproducible concentration estimates, and indicated that, on average, single mosquitoes yielded approximately 300 ng of DNA. Such a total yield is currently insufficient for many highthroughput genome screening applications, necessitating whole genome amplification of all or most individuals in a population prior to genotyping.展开更多
文摘Accurate estimates of DNA quantity are likely to become increasingly important for successful genomic screening of insect populations via recently developed, highly multiplexed genotyping assays and high-throughput sequencing methods. Here we show that genomic DNA extractions from single Anopheles gambiae Giles using a standard commercial kit-based methodology yield extracts with concentrations below the linear range of spectrophotometric absorbance at 260 nm. Concentrations determined by spectropho- tometry were not reproducible, and are therefore neither accurate nor reliable. However, DNA quantification using a fluorescent nucleic acid stain （PicoGreen） gave highly reproducible concentration estimates, and indicated that, on average, single mosquitoes yielded approximately 300 ng of DNA. Such a total yield is currently insufficient for many highthroughput genome screening applications, necessitating whole genome amplification of all or most individuals in a population prior to genotyping.
